Project Summary The activation and recruitment of brown adipose tissue has become an exciting target in the fight against obesity and its related metabolic diseases. Brown adipose tissue differs from white adipose tissue in its critical ability to burn energy as heat through UCP-1 driven adaptive thermogenesis. Expression of mitochondrial UCP-1 uncouples oxidative phosphorylation, resulting in accelerated substrate oxidation but low ATP production. The resulting dissipation of energy is associated with plasma triglyceride clearance, a key factor in weight loss. The brown adipose depot has long been known to exist in rodent models as well as in human infants. However, recent evidence has demonstrated the existence of active brown adipose tissue in adult humans. These findings prompted a resurgence of research focusing on both the activation of brown adipose tissue and the ?browning? of white adipose tissue, a phenomenon wherein UCP-1-mediated mitochondrial uncoupling is induced by stimuli. Multiple studies using rodent models have demonstrated that activation of brown fat or induction of browning can promote beneficial metabolic effects, and promising therapeutic targets have been identified. However, the therapeutic potential of these targets has not been confirmed due to the lack of a widely available human brown adipocyte model. Therefore, there is a pronounced need to validate existing therapeutic targets in the human system. ZenBio will address this need by providing a well characterized human brown adipocyte cell culture system to the research community and will utilize this tool to screen potential candidates for stimulation of human brown adipogenesis. The overarching goal of this project is to generate and characterize an adult human brown adipocyte cell system. Aim 1 is intended to build on our successful establishment of a human fetal brown adipocyte system in Phase 1 and use molecular and functional characterization to generate a widely available adult human brown adipocyte system. This includes establishing conditionally immortalized adult brown preadipocyte lines for use in cell-number intense research and screening. The second Aim will expand the adult system to include donors from a wide range of demographics. This Aim will also establish a database of molecular and functional characterization from this donor set that will be available to the research community. Aim 3 will establish a robust, high content/high throughput screening platform capable of multiplexing molecular and function al assays in a 384 well format. This platform will be able to screen for brown fat modulators and activators, as well as validating therapeutic targets. The goal of these three Aims is to characterize and commercialize the adult human brown adipocyte system from donors of differing BMI, age, gender and diabetic state and validate their use to identify novel targets of brown adipocyte differentiation for the treatment of obesity. These studies will result in innovative targets for human brown adipocyte activation, a crucial step for anti-obesity therapeutics.